Parkinson's disease (PD), PD with dementia (PDD) and dementia with Lewy bodies (DLB) are the the most common synucleinopathies. Although there is no cure for these diseases, passive immunization against ?- synuclein (?-syn) improves both the behavioral deficits and the neuropathology in mouse models of synucleinopathy and clinical trials testing the safety of monoclonal anti-?-syn antibodies in healthy volunteers and PD patients are ongoing. Despite the enthusiasm for ?-syn immunization, it is known that antibodies do no readily cross the blood brain barrier (BBB) and target engagement of ?-syn in the CNS may only be partial after the systemic antibody injection methods used in both rodent and human studies. Our hypothesis is that optimal neuroprotection may require higher anti-?-syn antibody accumulation in the brain. To overcome the challenge to immunization strategies for PD, PDD and DLB caused by the BBB, we will use low acoustic energy transcranial focused ultrasound stimulation (FUS), which opens the BBB reproducibly, selectively and safely, thereby enhancing antibody permeation to the CNS. Furthermore, FUS, in its own right, may mitigate ?-syn pathology. The rationale for the proposed research is that, once the anti-neurodegenerative effects of FUS alone and in combination of passive immunization in a mouse model of synucleinopathy are established, new and innovative strategies can be devised for the treatment of PD, PDD and DLB. So, the following two specific aims are proposed. In AIM 1, we will demonstrate the effects of FUS on behavior and neuropathology in a well-validated mouse model of synucleinopathy that will be injected with monoclonal anti-?-syn antibodies. Since mounting evidence indicates that microglia play a critical role in neurodegeneration and that FUS may modulate microglial function, in AIM 2, we will characterize the effects of FUS on the microglial response of a passively immunized mouse model of synucleinopathy. To achieve this stated goal, we will use state-of-the art single cell RNA sequencing technology to profile unbiasedly microglia from the brain of this mouse model of synucleinopathy that was subjected to FUS and/or immunization. In situ validation of the identified microglial subpopulations will also be performed in AIM 2. Successful completion of the proposed work is expected to overcome current limitations of using immunization for CNS disorders and will provide unique insights into the phenotypes of microglia in response to the disease process, FUS and/or immunization. These findings will have an important positive impact in that they will provide opportunities for therapeutic interventions and, fundamentally, advance our mechanistic understanding of PD and related disorders using ultrasound both for its potential direct effect on neurodegeneration and as an indirect immunotherapy enhancer through its action on the BBB.